Temporal lobe epilepsy is the most common form of epilepsy in adults and one of the most difficult types to treat. My long-range research goal is to contribute to a more complete understanding of the mechanisms that underlie this disorder. Previous studies suggest that in patients and models of temporal lobe epilepsy, layer III entorhinal cortical neurons die, and layer II neurons become hyperexcitable and deliver hypersynchronous excitatory synaptic input to the dentate gyrus. The goal of the proposed project is to identify the mechanisms of hyperexcitability and hypersynchrony in layer II of the entorhinal cortex of pilocarpine-induced epileptic rats. The specific aims are to: 1. Test the hypothesis that GABAergic interneurons are lost in the entorhinal cortex using in situ hybridization for glutamic acid decarboxylase, immunocytochemistry for somatostatin and parvalbumin, Nissl staining, and the optical fractionator method. 2. Test the hypothesis that layer II neurons receive less inhibitory synaptic input, because GABAergic interneurons are disconnected from their normal excitatory synaptic input. a. Inhibition of principal neurons in layer II of entorhinal cortical slices will be evaluated using whole-cell voltage-clamp recording of evoked, spontaneous, and miniature IPSCs. b. Spontaneous and miniature EPSCs in interneurons of pilocarpine-treated and control rats will be compared. 3. Test the hypothesis that layer II neurons receive excessive excitatory synaptic input. a. Excitatory synaptic input to principal neurons in layer II of the entorhinal cortex will be evaluated using whole-cell voltage-clamp recording of spontaneous and miniature EPSCs. b. Individual principal neurons in regions proposed as likely sources of new excitatory synaptic input to layer II (the presubiculum and layers II, V, and VI of entorhinal cortex) will be labeled with biocytin and 3-dimensionally reconstructed to measure their axon projections and determine if they sprout axon collaterals that may synapse with layer II neurons. c. Focal stimulation by glutamate photolysis will be used to compare the extent of monosynaptic excitatory input to principal neurons in layer II from the presubiculum and layers II, V, and VI of the entorhinal cortex.